CrkL activates integrin-mediated hematopoietic cell adhesion through the guanine nucleotide exchange factor C3G

CrkL is a member of the Crk family of adapter proteins consisting mostly of SH2 and SH3 domains, CrkL is most abundantly expressed in hematopoietic ce lls and has been implicated in pathogenesis of chronic myelogenous leukemia . However, its function has not been precisely defined. Here, we show that overexpression of CrkL enhances the adhesion of hematopoietic 32D cells to fibronectin. The CrkL-induced increase in cell adhesion was blocked by anti bodies against VLA-4 (alpha 4 beta 1) and VLA-5 ((alpha 5 beta 1) but was o bserved without changes in surface expression levels of these integrins. St udies using CrkL mutants demonstrated that the SH2 domain is partially requ ired for enhancing cell adhesion, whereas the C-terminal SH3 domain as well as the tyrosine phosphorylation site (Y207) is dispensable. In contrast, t he N-terminal SH3 domain, involved in binding C3G and other signaling molec ules, was showed to play a crucial role, because a mutant defective of this domain showed an inhibitory effect on the cell adhesion to fibronectin, Fu rthermore, overexpression of C3G also increased the adhesion of hematopoiet ic cells to fibronectin, whereas a C3G mutant lacking the guanine nucleotid e exchange domain abrogated the CrkL-induced increase in cell adhesion. On the other hand, a dominant negative mutant of H-Ras or that of Raf-1 enhanc ed the basal and CrkL-induced cell adhesion and that of R-Ras modestly decr eased the adhesion. Taken together, these results indicate that the CrkL-C3 G complex activates VLA-4 and VLA-5 in hematopoietic cells, possibly by act ivating the small GTP binding proteins, including R-Ras, through the guanin e nucleotide exchange activity of C3G. (C) 1999 by The American Society of Hematology.